The proliferation and differentiation of cells in multicellular organisms is subject to a highly regulated process. A distinguishing feature of cancer cells is the absence of control over this process; proliferation and differentiation become deregulated resulting in uncontrolled growth. Significant research efforts have been directed toward better understanding this difference between normal and tumor cells. One area of research focus is growth factors and, more specifically, autocrine growth stimulation.
Growth factors are polypeptides which carry messages to cells concerning growth, differentiation, migration and gene expression. Typically, growth factors are produced in one cell and act on another cell to stimulate proliferation. However, certain malignant cells, in culture, demonstrate a greater or absolute reliance on an autocrine growth mechanism. Malignant cells which observe this autocrine behavior circumvent the regulation of growth factor production by other cells and are therefore unregulated in their growth.
Study of autocrine growth control advances understanding of cell growth mechanisms and leads to important advances in the diagnosis and treatment of cancer. Toward this end, a number of growth factors have been studied, including insulin-like growth factors (“IGF-I” and “IGF-II”), gastrin-releasing peptide (“GRP”), transforming growth factors alpha and beta (“TGF-a” and “TGF-b”), and epidermal growth factor (“EGF”).
The present invention is directed to a recently discovered growth factor. This growth factor was first discovered in the culture medium of a highly tumorigenic “PC cell line,” an insulin-independent variant isolated from the teratoma derived adipogenic cell line 1246. This growth factor is referred to herein as “GP88.” GP88 has been purified and structurally characterized. Amino acid sequencing of GP88 indicates that GP88 has amino acid sequence similarities with the mouse granulin/epithelin precursor.
Granulins/epithelins (“grn/epi”) are 6 kDa polypeptides and belong to a novel family of double cysteine rich polypeptides. U.S. Pat. No. 5,416,192 (Shoyab et al.) is directed to 6 kDa epithelins, particularly epithelin 1 and epithelin 2. According to Shoyab, both epithelins are encoded by a common 63.5 kDa precursor, which is processed into smaller forms as soon as it is synthesized, so that the only natural products found in biological samples are the 6 kDa forms. Shoyab et al. teaches that the epithelin precursor is biologically inactive.
Contrary to the teachings of Shoyab et al., the inventor's laboratory has demonstrated that the precursor is not processed as soon as it is synthesized. Studies, conducted in part by this inventor, have demonstrated that the precursor (i.e., GP88) is in fact secreted as an 88 kDa glycoprotein with an N-linked carbohydrate moiety of 20 kDa. Analysis of the N-terminal sequence of GP88 indicates that GP88 starts at amino acid 17 of the grn/epi precursor, demonstrating that the first 17 amino acids from the protein sequence deduced from the precursor cDNA correspond to a signal peptide compatible with targeting for membrane localization or for secretion.
Also in contrast to the teachings of Shoyab et al., the inventor demonstrated that GP88 is biologically active and has growth promoting activity, particularly as an autocrine growth factor for the producer cells.
Hematopoietic malignancies are malignant blood diseases including various lymphomas and leukemias. Leukemias of B-cell lineage include, but are not limited to, acute lymphocytic leukemia, B cell lymphoma, and multiple myeloma. Multiple myeloma (“MM”) is a clonal B-cell neoplasm and the second most prevalent blood cancer, representing 1% of all cancers and 2% of all cancer deaths. B-cells (or B-lymphocytes) are precursor cells that differentiate into plasma cells after exposure to particular antigens. Plasma cells produce immunoglobulins and have a limited life span. However, uncontrolled growth of plasma cells in a clonal lineage of B cells may lead to accumulation of plasma cells producing monoclonal immunoglobulins or immunoglobulin fragments (e.g., M protein). MM is characterized by bone degradation and fractures, anemia, increased risk of infection, and decreased production of platelets in addition to other symptoms. The incidence of MM, currently about 14,000 new cases per year, has been steadily increasing in the United States for several decades (1). There has been little improvement in the treatment of human MM over the past 25 years and there is no cure for the disease (3). The few available therapies for treatment of MM have severe side effects and are of limited efficacy. For nearly 3 decades, the standard treatment for human MM has been glucocorticoid and/or chemotherapy with melphalan and prednisone alone or combinations of alkylating agents such as glucocorticoids and anthracyclines (4). However, almost all patients with MM who initially respond to glucocorticoid therapy relapse, with a median survival of two to three years following diagnosis (5). During the progression of MM to more aggressive forms of the disease, MM cells become insensitive to the killing effect of glucocorticoids leaving only the use of chemotherapeutic agents to control the disease.
What is needed are new compositions and methods for treatment and diagnosis of MM, and particularly compositions and methods that inhibit the proliferation and survival of multiple myeloma cells.